POS1014 INVESTIGATING THE SYNOVIAL PATHOLOGY RELATED TO TREATMENT RESISTANCE IN JAPANESE RHEUMATOID ARTHRITIS PATIENTS USING SINGLE-CELL ANALYSIS
Authors: R. Yohihara, H. Tsuchiya, Y. Omata, H. Takahashi, S. Nakajima, H. Mizuhara, A. Uchio, Y. Maenohara, Y. Fujieda, T. Matsumoto, H. Harada, H. Shoda, N. Tamura, T. Mimura, K. Ohmura, A. Morinobu, T. Atsumi, Y. Tanaka, T. Takeuchi, S. Tanaka, T. Okamura, K. Fujio
Journal: Annals of the Rheumatic Diseases (2023)

<h3>Background</h3> Despite of recent developments in therapeutic agents for rheumatoid arthritis (RA) patients<sup>[1]</sup>, it is reported that half of the patients are unable to achieve remission even with existing drugs<sup>[2]</sup>. Therefore, there is an urgent need to gain mechanistic insight into treatment resistance. Nowadays, single-cell RNA sequencing (scRNA-seq) technology have dramatically improved our understanding of the heterogeneity in synovial cells. However, it has not been fully elucidated how the cell clusters are related to treatment response, especially in Asian races<sup>[3]</sup>. <h3>Objectives</h3> We intend to analyze the RA synovium from Japanese patients based on single-cell transcriptomics to explore the pathological key players related to treatment resistance. <h3>Methods</h3> Synovial specimens were collected from 31 RA patients using an ultrasound-guided needle biopsy. The proportion of 5 immune cell subsets (CD4<sup>+</sup> T cells, CD8<sup>+</sup> T cells, B cells, NK cells, monocytes) and mesenchymal cells (synovial fibroblasts (SF), endothelial cells, mural cells) were analyzed by flow cytometry. CD45<sup>+</sup> and CD45<sup>-</sup> live cells were isolated, and scRNA-seq libraries were prepared using the 10x chromium system. <h3>Results</h3> We classified the patients into the following three groups based on treatment status at the time of biopsy; treatment-naïve, inadequate response to conventional synthetic disease-modifying antirheumatic drugs (csDMARDs-IR), or biological DMARDs (bDMARDs-IR). The proportion of CD8<sup>+</sup> T cells, especially <i>GZMB</i><sup>+</sup><i>GZMK</i><sup>+</sup> CD8<sup>+</sup> T cells, was significantly lower in csDMARDs-IR patients compared to treatment-naïve patients. This population was characterized by enhanced expression of <i>IFNG</i> and <i>GZMK</i>, the cooperative inducers of IL-6 production from SF. Meanwhile, an increased proportion of SF, especially <i>THY1</i><sup>low</sup> sublining and <i>CD34</i><sup>+</sup> sublining, was observed in csDMARDs-IR and bDMARDs-IR patients. Intriguingly, by integrating gene set variation analysis (GSVA) with transcriptomic data of cytokine-stimulated SF <i>in vitro</i><sup>4</sup>, <i>THY1</i><sup>low</sup> sublining was indicated to be activated independently of the effects of inflammatory cytokines (e.g., TNF-α, IL-1β, IFNs) (Figure 1). Collectively, this SF subpopulation was inferred to be less susceptible to cytokine-blocking agents such as IL-6 receptor or TNF-α inhibitors. <h3>Conclusion</h3> The synovial analysis has the potential to be useful in parsing the mechanism of treatment resistance in Japanese RA patients, and gaining insights into novel therapeutic targets. <h3>References</h3> [1] Tan Y, Buch MH. 'Difficult to treat' rheumatoid arthritis: current position and considerations for next steps. RMD Open. 2022 Jul;8(2):e002387. [2] Burmester GR, Bijlsma JWJ, Cutolo M, et al. Managing rheumatic and musculoskeletal diseases - past, present and future. Nat Rev Rheumatol. 2017 Jul;13(7):443-448. [3] Chang F, Wei K, Slowikowski K, et al. Defining inflammatory cell states in rheumatoid arthritis joint synovial tissues by integrating single-cell transcriptomics and mass cytometry. Nat Immunol. 2019;20(7):928-42. [4] Tsuchiya H, Ota M, Sumitomo S, et al. Parsing multiomics landscape of activated synovial fibroblasts highlights drug targets linked to genetic risk of rheumatoid arthritis. Ann Rheum Dis. 2021 Apr;80(4):440-450. <h3>Acknowledgements:</h3> NIL. <h3>Disclosure of Interests</h3> None Declared.